Take These Genes and Call Me in the Morning

In a concrete bunker at Fort Detrick Army Base in Frederick, Maryland, Jenny Riemenschneider is standing over 10 rabbits splayed out on stainless steel operating tables. Dressed in a white Tyvek jumpsuit, a surgical mask, a shower cap, and plastic booties, she calmly loads 12 gold bullets into a revolver. Each rabbit lies motionless, tranquilized, paws spread-eagle with an exposed patch of shaved skin on its lower abdomen. Riemenschneider grips the pistol with both hands and presses its 8-inch barrel into a pink belly. Boof! Boof! Boof! Boof! Boof! Boof! Boof! Boof! She fires eight shots into the first rabbit, flinching slightly as the blasts echo through the room.

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A smile rises from behind her mask.

Nigel CoxBio-Rad's gene gun:
1. A blast of helium fires tiny gold pellets carrying harmless snippets of a pathogen's DNA into the skin.
2. The DNA infiltrate the nucleus of the skin and muscle cells.
3. The DNA forces the cell to produce pathogen proteins, triggering the immune system.
4. The immune system unleashes killer T cells to attack foreign proteins and learns how to deal with the actual virus.

"Looks good," she says. "See those clouds — those faint blushes just beneath the skin? Those are the bullets. It's a perfectly painless procedure."

Riemenschneider is part of a team of military scientists experimenting with so-called gene vaccines as a weapon against the growing threat of bioterror. Her revolver? A medical device known as a gene gun, which fires capsules containing thousands of DNA-coated gold pellets designed to inoculate the bunnies against anthrax. Propelled into the rabbit's skin cells by a blast of compressed helium, the DNA fragments are supposed to train the animal's immune system to recognize and combat the actual disease. Six months later, when Riemenschneider exposes the rabbits to what should be a deadly dose of anthrax, she's ready to declare success: Nine out of 10 remain healthy.

Gene vaccines may be relatively new, but they're the logical outgrowth of two familiar strands of medical science. First is the 200-year-old practice of vaccination, in which the body is infected with a weakened form of a disease that prepares the immune system for a future encounter with the real thing. Traditional vaccines are highly effective at conferring long-term immunity against diseases like measles, mumps, and polio, but because they involve growing and injecting a live pathogen, they're costly, cumbersome to produce and transport, and too dangerous to use against super-virulent viruses like HIV. What's more, traditional vaccines are effective only against infectious diseases — afflictions like cancer and Alzheimer's are left to more radical treatments, such as chemotherapy and surgery.

While immunologists grappled with such limitations in the 1970s, an explosion of knowledge in the field of genetics led to a new approach to tackling disease: gene therapy. Gene therapy aims to conquer genetic diseases by replacing targeted genes. The concept was promising, but the medical record has been unsuccessful because the body's immune system rejects therapeutic DNA as foreign — just as it would reject a common bug.

Gene vaccines borrow from both traditional vaccinology and gene therapy. By isolating a harmless snippet of a pathogen's DNA and injecting it into the body, researchers believe they can fool the immune system into developing an attack plan against a particular disease even though the body was never exposed to it. Whereas gene therapy tries to work in spite of the immune system, gene vaccines harness the immune system's instinct to search out and destroy alien proteins. "I still can't believe it actually works," says Riemenschneider, who has spent seven years investigating killer viruses such as Ebola. "DNA vaccines are incredibly easy to make. You can produce them in days or weeks, whereas the traditional methods often take years."

Gene vaccines hold special promise as weapons against diseases too complex or dangerous for traditional immunology. Already, they've proven successful in hundreds of animal trials against bioweapons like anthrax and the plague, as well as against pandemics like malaria and TB, which claim millions of lives each year. In July, Oxford scientist Adrian Hill began testing a gene-based malaria vaccine on hundreds of at-risk people in Gambia.

Closer to home, a gene vaccine against melanoma has completed three rounds of clinical trials on humans and appears ready to be submitted to the FDA for final approval. When injected directly into cancerous tumors, the vaccine, called Allovectin-7, causes proteins to grow on the tumor's surface — which in turn stimulates the immune system. The drug's manufacturer, Vical, is reviewing data from the experiments in hopes of presenting them to the FDA. If the drug gets a thumbs-up, Allovectin-7 may be on the market as soon as next year — and may unleash a torrent of new research. "When the flagship product makes it through the process, it will be a landmark proof of principle," says Vijay Samant, Vical's president. "Investment dollars will pour into both the vaccine and gene therapy industries."

The same principle that allows gene vaccines to destroy melanoma is being applied to diseases once thought resistant to immunization. In April, Merck announced that its gene-based HIV vaccine had induced immunity in more than half of the 300 human subjects in its ongoing phase 1 trial. These results, by far the most successful yet for an AIDS vaccine, astonished the medical community. "This is without question the most promising technology that has come along for an AIDS vaccine," says Jeffrey Laurence, senior scientific consultant at the American Foundation for AIDS Research, "but keep in mind that we still have a long, long way to go to find a cure."

The distinction between inducing immunity and preventing infection is critical: While the Merck vaccine successfully bolstered immune response — significantly slowing the infection process and reducing the likelihood of full-blown AIDS — it did not prevent infection altogether. Any gene-based HIV vaccine will require years of research before it can officially be proven effective and brought to market.

Meantime, there are vaccines in the pipeline for bacterial diseases like anthrax, viral pathogens like Ebola, and inheritable diseases, including several forms of cancer and Alzheimer's. An Alzheimer's vaccine, for example, would stimulate the immune system to attack the protein deposits in the brain that are caused by the degenerative disorder. The same principle could be applied to all sorts of health issues. There's even talk of gene vaccines being used to prevent pregnancy (by training the immune system to attack the cells that produce sperm) and to conquer drug addiction (by blocking the brain's receptivity to the drug). But why stop there? "There's evidence that the elimination of a particular glucose response element in the cell makes mice live longer. We could selectively eliminate that receptor by immunization," says Stephen Albert Johnston, director of the Center for Biomedical Inventions at the University of Texas Southwestern Medical Center and a leader in gene vaccine research. "This technology has transformed our understanding of what a vaccine can do. Not just to prevent disease, but to probe the complex strategies of the immune system so we can use them to our advantage."

Traditional vaccines date back to the 1790s. With a smallpox epidemic plaguing the British population, physician Edward Jenner noticed that milkmaids were the only people with pock-free skin and reasoned that their exposure to cowpox, a less virulent disease, had conferred immunity. He tested his theory by filling a cut on an 8-year-old boy's arm with liquid from a cowpox pustule. A few months later, he took his experiment to a sublime end: He repeated the procedure, this time using the ooze from a smallpox pustule. The boy, Jenner discovered, was immune.

Listen to Maurice Hilleman, who invented the standard vaccines for measles, influenza, and chicken pox while at Merck, and he'll tell you the basic principles of vaccinology advanced very little until the 1970s. That's when he and some colleagues discovered the immune system could learn to combat a pathogen from a telltale selection of its proteins. "It's like giving a bloodhound a whiff of a criminal's clothes before the hunt," says Hilleman. In 1986, the FDA approved the first vaccine of this kind, made of lab-grown recombinant proteins, for hepatitis B. In the early '90s, three scientists — Jon Wolff at the University of Wisconsin, UT Southwestern's Johnston, and Margaret Liu at Merck — made independent discoveries revealing that pure DNA could be a simpler and dramatically more effective medium.

The basic science is simple. Everything in the body — from bones to hormones — is made of proteins. DNA provides the instructions for producing proteins; cells crank them out. To replicate, a virus must penetrate a host's cell and insert its own genetic material, forcing the cell to manufacture more copies of the virus. Our immune system fights this invasion with a web of specialized sentinel cells, spread throughout flesh, muscles, and organs, which scan every protein in the body. "When the sentinel cells get activated," says Hilleman, fluttering his fingers, "they scootle over to the nearest lymph nodes with a message: 'We've been invaded! Mobilize the troops.'" Immune cells produce antibodies that then try to eliminate the disease in the bloodstream. But if the intruder blows past and begins to replicate, the system mobilizes killer T cells to search out and destroy infected cells, and creates a reserve batch of T cells to eliminate that type of intruder in the future.

Traditional vaccines force the body to create reserve killer T cells by infecting it with a mild form of a disease. Some viruses are too dangerous to be injected live, because even in a weakened state, they can outwit the immune system. HIV camouflages itself to elude surveillance and infects the body so furtively that the immune system can't react in time. But even HIV reveals its defining proteins when it invades a cell — precisely the information that a gene vaccine can train the body to detect.

The problem is not isolating those proteins. What's needed is a delivery mechanism that inserts the protein-producing genes into enough of the body's cells to stimulate an enduring immune response. Scientists first experimented with a bug like the common cold, replacing the contagion's content with the desired DNA. After all, viruses have evolved over millions of years with the sole purpose of fully infiltrating a host, and they are incredibly efficient at doing so. Why not, then, use that expertise as a vehicle for the genes? The good news about viral vectors, as they're called, is that they cost only about $10 a dose, compared with $40 for a traditional vaccine. The bad news is that, like traditional vaccines, they require the cultivation and transportation of live viruses.

One way to get around this problem is to inject the DNA directly into the body with a needle or a gun — no virus necessary — as Riemenschneider did with the rabbits. Once the DNA breaks through the skin, it infiltrates the nucleus of cells near the surface, much like a viral vector would, forcing the cell to manufacture the pathogen's proteins. It's a process that has been patented by Vical. So-called naked DNA is more stable, is easier to manufacture and transport than traditional vaccines or viral vectors (live viruses require refrigeration), has fewer health risks, and costs only about 40 cents per dose. What's more, the body cannot develop immunity to DNA the way it can to a virus carrying a gene vaccine.

Of course, naked DNA is nowhere near as efficient as viruses are, so scientists are scrambling to develop more powerful ways of blasting it into the body. British biotech firm PowderJect upgraded its gene gun to use smaller particles and a stronger blast of helium — while also streamlining the design and adding a silencer. Another gene gun manufacturer, Bio-Rad, uses an adjustable low-pressure helium pulse. The San Diego company Genetronics has perfected a procedure called electroporation which uses electrical fields to open pores in the cell membranes to clear the way for naked DNA. "If naked DNA technology can be optimized, it will be — and I mean this literally — the ultimate killer app," says Margaret Liu, who left Merck and now is an adviser to the Gates Foundation on its vaccination efforts. Liu says the simplicity of gene vaccines would make them attractive to developing countries.

Some more far-out gene delivery methods are also in the works, from nasal sprays and transdermal patches to bioengineered fruits and vegetables. Hugh Mason at Cornell University has reengineered potatoes with gene vaccines for both hepatitis B and human papilloma virus — the main cause of cervical cancer. The desired proteins are spliced into the seed and appear in the plant flesh; when consumed, they trigger sentinel cells in the stomach lining. Robert Webb at the US Army Medical Research Institute for Infectious Diseases bioengineered a plague vaccine into tomatoes. Scientists are also engineering vaccines for rotavirus, Norwalk virus, and tooth decay into bananas, corn, and apples.

The near-term prospects of gene vaccines are about to be put in the hands of the FDA, when Vical submits Allovectin-7 for review. The scientific community will be watching closely. "The first DNA vaccine to make it through will be pathfinding," says David Baltimore, the Nobel Prize-winning president of Caltech. "Eventually these products will happen."